Quality Assurance for Pneumococcal Assays in Europe

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Quality Assurance for Pneumococcal Assays in
Europe

• Daniel Harrison

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Index

• Background to the study
• Pneumococcal pathogen
• Pilot pneumococcal serotype study

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A European QA Scheme for Pneumococcal Assays

• EU funded Consensus conference (Langen 2006) on
  Primary Immunodeficiency Diseases (PIDs)
  outlined
• A lack of quality assurance for the assays that
  are used for measuring specific antibody assays
  to common pathogens and immunization antigens
  within PID community!

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A European QA Scheme for pneumococcal assays

• These assays are an important aid for the
  diagnosis of primary antibody deficiencies.
• Provision of standardized assay protocols with a
  reliable quality assurance scheme will enable
  progress in determinating diagnostic criteria for
  these diseases.
• For this reason A study to develop a quality
  assurance scheme for pneumococcal assays in
  Europe was initiated.

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ESID guidelines

• Currently pneumovax unconjugated vaccine is
  included in the ESID guidelines for PIDs. E. De
  Vries 2005
• Is used to determine the severity of humoral
  immunodeficiency in a patient by way of looking
  at the result from booster vaccinations
• However the authors recognise that the protocols
  will need to be revised
• Pneumococcal serotype assays could be
  incorporated

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The Pathogen - Streptococcus pneumoniae

• What Is it?
• Common microflora bacteria that has gt90 known
  serotypes
• Because of this variation it has strong
  pathogenicity

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Streptococcus pneumoniae

• What does it cause?
• Causes pneumonia, meningitis and systemic sepsis.
  
• Prevalent in individuals at the extremes of age
• 1 million deaths are accountable to pneumococcal
  infections worldwide
• This is mostly in developing countries

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PIDs

• Are highly susceptible to infections caused by
  encapsulated bacteria
• Especially those with X- linked
  agammaglobulinaemia and common variable immune
  deficiency

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Invasive Pneumococcal Disease

• What is it and what are the symptoms?
• The most severe form of pneumococcal infection
• Three major conditions
• pneumonia, bacteremia, and meningitis.
• They are all caused by infection with the same
  bacteria, but have different symptoms.

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Vaccines

• 2 available pneumococcal vaccines pneumococcal
  polysaccharide vaccine (PPV) or Pneumovax and
  the pneumococcal conjugate vaccine (PCV) or
  Prevenar
• Pneumovax contains chains of polysaccharides
  that make up the surface capsule of the bacteria.
  
• The 23 types of pneumococci that are included
  cause 88 of invasive pneumococcal disease.
• This vaccine is not effective in children younger
  than 2 years in whom 80 of invasive pneumococcal
  disease in childhood occurs so it is often
  given to elderly patients

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Vaccines and Serotype Prevalence in USA and
Europe

• Prevenar includes purified capsular
  polysaccharide of seven types of the bacteria
  conjugated to a harmless variety of diphtheria
  toxin.
• The seven types 4, 6B, 9V, 14, 18C, 19F and 23F
  account for 86 of bacteremia, 83 of meningitis,
  and 65 of acute otitis media among children less
  than six years of age in the United States.
• These seven serotypes are responsible for 83 of
  invasive pneumococcal disease in children lt4
  years old in the US and cause the majority of
  invasive disease in Europe
• All healthy infants and toddler should receive
  four doses of Prevenar vaccine

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Pilot Study

• Aims
• To develop a quality assurance scheme for
  pneumococcal assays in Europe that aid the
  diagnosis of PIDs.
• To compare pneumococcal serotype results with
  those from the existing 23 valent Pneumovax ELISA
  assay, in order to determine the relative
  usefulness of these assays for test immunisation
  in the diagnosis of PIDs
• To determine if it is possible to establish
  levels of protection in patients at high risk of
  invasive pneumococcal disease

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Pilot Study

• Methods
• Development of ELISA assays, in accordance with
  WHO protocol, for the detection of IgG reactive
  to 7 common pneumococcal serotypes (4, 6B, 9V,
  14, 18C, 19F and 23F) in human serum
• Acquired and tested 16 control serum samples
  along with 16 known positive pneumovax serum
  samples
• Concentrations of specific IgG to certain
  serotypes, within sera, were calculated using the
  reference serum from WHO (89-SF)

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Protection Level

• Needed to have a protection level in order to
  establish positive and negative results
• Black et al. 2000 found that gt95 of patients
  receiving the pneumococcal conjugate vaccine
  developed gt0.15 µg/ml after 3rd dose
• Henckaerts et al. 2005 found that the putative
  protection level was higher (3.5) in the non 22F
  assay
• Therefore 0.2 µg/ml was used as a putative
  protection level

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Results

• Wide range between controls, but also
  between serotypes

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Serotype Specific Immunoglobulin G
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Statistical Correlation with 7 serotypes and
Pneumovax Assay(Kappa Coefficients)
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Pilot study

• Next Steps
• Samples with adequate volumes and concentrations
  will be selected for distribution to 15 centres
  in Europe
• This is so that our assay can be compared and
  validated with those of our potential European
  collaborators

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Pilot study

• Next Steps
• Will attempt to make a selection of sera
  including 2 with high concentrations of specific
  anti-pneumococcal antibodies, 2 with low and 2
  with intermediate concentrations
• There are anomalies, but we will inform
  collaborators of these

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Conclusions

• Observed high immunity for serotypes 14 and 18C
• Found that immunity against the serotypes varies
  between serotypes and individuals
• Immunity against most serotypes generally
  indicates a high pnuemovax result but not always
• Compared serotype findings to results obtained
  from the existing total Pneumovax ELISA assay and
  found a correlation with 23F serotype

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Acknowledgements

• Dr Helen Chapel
• Dr Berne Ferry